Cloning and sequencing of chiC gene of Bacillus circulans WL-12 and relationship of its product to some other chitinases and chitinase-like proteins

Abstract

To clarify the roles of individual chitinases comprising in the chitinase system of Bacillus circulans WL-12, the gene ( chiC) encoding chitinase C, a minor constituent of the chitinase system, was cloned and expressed in Escherichia coli. The predicted product of the chiC gene is 491 amino acids long with a calculated molecular mass of 53,447 Da. The N-terminal portion of the deduced polypeptide exhibited 46.8% amino acid homology with the catalytic domain of chitinase A1 of this bacterium, suggesting that the chitinase encoded by the chiC gene is comprised of an N-terminal catalytic domain and a C-terminal domain with unknown function. Two bands of chitinases with estimated molecular masses of 55 kDa and 40 kDa were detected on SDS-PAGE of the periplasmic fraction of E. coli carrying the cloned chiC gene. Chitinase C of B. circulans WL-12 is practically identical to the 40 kDa chitinase detected in E. coli based on their N-terminal amino acid sequences, isoelectric points and estimated sizes, and corresponds to the catalytic domain of the initial product of the chiC gene (55 kDa chitinase, designated as chitinase C1). Comparison of domain organizations of chitinases, of this bacterium, sequenced so far suggested that they represent two types of catalytic domains and that domain shuffling occurred relatively recently giving rise to three chitinases. Sequence comparison and the evolutionary relationship of chitinase C1 with other chitinases and chitinase-like proteins are also discussed.